Listeria monoctyogenes is a facultative intracellular human pathogen and a model organism with which to explore basic mechanisms of pathogenesis and host defense. This research proposed in this application relies heavily on the use of bacterial and host mutants to examine a macrophage cytosolic surveillance pathway of innate immunity that detects intracellular pathogens. Activation of the cytosolic response leads to the expression of IFN-p and co-regulated genes. In Aim I,forward genetic screens will be used to identify L monocytogenes molecular determinants that contribute to stimulating the host cytosolic host pathway of innate immune recognition. The screen has been validated by the identification and characterization of a bacterial multidrug resistance transporter (MDR) required to activate the host cytosolic pathway. In Aim II, the genetic results in Aim I will guide studies to identify the bacterial ligand(s) and cognate host receptor(s). The precise identity of the ligands will be determined using analytical tools and take advantage of a pair of strains that vary 60-fold in their induction of the host cytosolic surveillance pathway. In Aim III,the relevance of the bacterial determinants that govern the host innate immune response will be evaluated by examining mutants in four animal models of infection including: the systemic murine listeriosis model in naive and immunized mice, a pregnant Guinea Pig model that monitors trafficking to the placenta, a newly developed oral model of listeriosis in immunocompromised mice that leads to infection of the brain, and finally, the capacity to induce immunity to L. monocytogenes and foreign antigens. The last model has relevance to vaccine development. If successful, these studies will lead to the discovery of a novel microbial molecule that stimulates a host pathway that contributes to the development of innate and acquired immunity. RELEVANCE (See instructions): The proposed studies on Listeria monocytogenes will provide insight into the pathogenic mechanisms of a lethal food-borne pathogen that causes infection of the brain and miscarriages. Further, these studies will lead to the characterization of a host system of innate immunity that will lead to vaccines and/or therapeutics to treat disease, with relevance to biodefense, emerging infections and global health. PROJECT/PERFORMANCE S1TE(